Staphylococcus aureus is emerging as the most problemafic bacterial pathogen facing our community and healthcare settings. An effective strategy for S. aureus to survive in the host is to attach to a surface and develop into an encased community of cells called a biofilm. We recently discovered that quorum-sensing can control the balance between a planktonic or biofilm lifestyle, suggesting that modulation of this dispersal mechanism could be an effective therapeutic strategy. In collaboration with Dr. Kenneth Bayles (the PI of this PPG), we demonstrated that a deletion of the S. aureus secreted nuclease (Nuc) caused an overall thickening of the biofilm and inhibited secondary structure formation, and we have confirmed a recent report that S. aureus possesses a second extracellular nuclease activity (Nuc2). Based on these findings, our central hypothesis is that control over extracellular nuclease activity is a critical determinant of biofilm maturation and dispersal. To address this question, in Specific Aim 1 we will (i) define the role of Nuc and Nuc2 in biofilm maturation;(ii) determine whether nuclease activity is important for biofilm dispersal;and (iii) modulate biofilm integrity with controlled exposure to nuclease. We further propose that S. aureus nuclease is an important virulence factor. To investigate the nuclease function in disease, we will work with Dr. Tammy Kielian (Project 4 leader) and (i) examine the role of nuclease activity in evasion of neutrophil extracellular traps (NETs);(ii) define the significance of nuclease in mouse models of planktonic versus biofilm infection;and (iii) compare the host inflammatory response to nuclease in planktonic versus biofilm infection. Finally, we speculate that small-molecule inducers of nuclease activity could serve as anti-biofilm therapeutics. Towards this end, in Specific Aim 3, we will employ new technology to generate cyclic peptide libraries in S. aureus that are amenable to high-throughput screening methods. More specifically, we will (i) screen for cyclic peptides that induce nuclease expression through FACS;(ii) perform molecular and biochemical studies to identify peptide targets;(iii) characterize the best candidates as dispersal agents in a biofilm infection model;and (iv) compare results to transposon mutants with increased nuclease activity. Overall, the goal of this Project is to understand how these S. aureus biofilm structures form and disassemble, the contribution of extracellular DNA to this process, and the relevance in disease.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Research Program Projects (P01)
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Special Emphasis Panel (ZAI1-TS-M)
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University of Nebraska Medical Center
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Yamada, Kelsey J; Kielian, Tammy (2018) Biofilm-Leukocyte Cross-Talk: Impact on Immune Polarization and Immunometabolism. J Innate Immun :1-9
Bhinderwala, Fatema; Lonergan, Samantha; Woods, Jade et al. (2018) Expanding the Coverage of the Metabolome with Nitrogen-Based NMR. Anal Chem 90:4521-4528
Heim, Cortney E; Vidlak, Debbie; Odvody, Jessica et al. (2018) Human prosthetic joint infections are associated with myeloid-derived suppressor cells (MDSCs): Implications for infection persistence. J Orthop Res 36:1605-1613
Svechkarev, Denis; Sadykov, Marat R; Bayles, Kenneth W et al. (2018) Ratiometric Fluorescent Sensor Array as a Versatile Tool for Bacterial Pathogen Identification and Analysis. ACS Sens 3:700-708
Yamada, Kelsey J; Heim, Cortney E; Aldrich, Amy L et al. (2018) Arginase-1 Expression in Myeloid Cells Regulates Staphylococcus aureus Planktonic but Not Biofilm Infection. Infect Immun 86:
King, Alyssa N; Borkar, Samiksha; Samuels, David J et al. (2018) Guanine limitation results in CodY-dependent and -independent alteration of Staphylococcus aureus physiology and gene expression. J Bacteriol :
Mlynek, Kevin D; Sause, William E; Moormeier, Derek E et al. (2018) Nutritional Regulation of the Sae Two-Component System by CodY in Staphylococcus aureus. J Bacteriol 200:
Gries, Casey M; Kielian, Tammy (2017) Staphylococcal Biofilms and Immune Polarization During Prosthetic Joint Infection. J Am Acad Orthop Surg 25 Suppl 1:S20-S24
Krute, Christina N; Rice, Kelly C; Bose, Jeffrey L (2017) VfrB Is a Key Activator of the Staphylococcus aureus SaeRS Two-Component System. J Bacteriol 199:
Moormeier, Derek E; Bayles, Kenneth W (2017) Staphylococcus aureus biofilm: a complex developmental organism. Mol Microbiol 104:365-376

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